This Program, consisting of four Projects and three Cores, focuses on parenchymal (intracerebral) arterioles, which distribute blood within the brain and exhibit unique, albeit poorly understood, properties. The overall goal is to elucidate the molecular mechanisms by which the endothelium and smooth muscle (SM) of parenchymal arterioles (PAs) regulate vascular tone and neurovascular coupling (NVC) normally, and following ischemia/reperfusion (l/R) injury and subarachnoid hemorrhage (SAH). The central unifying theme is that the precise control of Ca2+ signaling in cells composing the "cerebrovascular unit"- endothelium, SM and astrocytes-determines normal brain function;by extension, dysfunction of Ca2+ signaling dynamics contributes to cerebrovascular disorders. Each project studies overlapping elements of the biological system to form a whole. Project 1 will provide the first information on properties and function of PA endothelial cells (ECs), exploring Ca2+ signaling and Ca2+-sensitive SK and IK potassium channels and their impact on SM function and NVC. Project 2 will focus on parenchymal arteriolar SM, with an emphasis on voltage-dependent Ca2+ channels (Cav) and transient receptor potential (TRP) channels. Projects 1 and 2 provide the platform for Projects 3 and 4. Project 3 will interact with Projects 1 and 2 to explore the underlying mechanistic basis for the profound changes in parenchymal arteriolar reactivity that develop following I/R, focusing on EC-SM changes in ion channel functionality. Project 4, which focuses on PAs and NVC following SAH, intersects with Project 2 on the effects of SAH on SM Cav and TRP channels, and with Project 1 on endothelial function and NVC. Because the endothelium and SM function physiologically as one entity, Projects 1 and 2 are inherently highly interdependent. Projects 3 and 4, which focus on dysregulation of cerebrovascular unit function under two divergent, but clinically relevant, pathological conditions, depend on information derived from Projects 1 and 2. The Imaging Core will provide state-of-the-art approaches for Ca2+ measurements in complex systems. The Animal and Instrumentation Core will consolidate and coordinate all animal and in vivo work, including the development of novel genetically encoded ratiometric Ca2+ biosensors. The long-term objective is to understand blood flow in the brain in health and disease, and by doing so, to reveal exciting novel targets that can be exploited in the treatment of cerebrovascular disease.